In this grant the major goal is to investigate mechanisms regulating quiescent B cells either poised in the GO stage of the cell cycle (GO B cells) or leaving GO to enter the cell cycle. We have found that caspases- a family of cysteine proteases involved in programmed death of B cells- also regulates B cell entry into the cell cycle. The caspase-6 inhibitor VEID blocks proliferation of GO B cells. However, caspase inhibitors do not block induction of inhibitors of apoptosis (lAPs). Our data suggest caspases are required for a pathway common to all modes of inducing B cells into the cell cycle. Our Aims are: Aim 1: To define the requirements for caspase activation in B cells, i.e., the signaling pathways upstream of caspase activation in B cells. We will test whether or not different receptors activate the same or distinct sets of caspases in GO B cells. We will test the hypothesis that T cell dependent (TD) vs. T cell independent (TI) signaling receptors activate similar or distinct caspase pathways in GO B cells. We will define the upstream pathways required for activating caspase-6 and -8 in B cells. Using a dominant negative (DN) form of FADD or caspase-6 and B cells from caspase-6 knockout mice, we will test if activation of caspase-6 is required for activation of caspase-8 or vice versa. We will also test if clAPs, when activated via CD40, contribute to the selective activation of caspases -6 and -8 and ceil cycle entry. Aim 2. We will test the hypothesis that caspase-6 cleaves a functional set of substrates in GO B cells as they enter the cell cycle and activate a survival program. First, we will characterize the cell cycle block observed upon inhibition of caspase-6. Second, we will define substrates for caspase-6 and assess their roles in B cells. We will define where caspase-6 goes in B cells after they are activated and how the caspase-6 substrate, SATB1, is regulated by caspase-6 in B cells. Aim 3. We will characterize the B cell defects in caspase-6 -/- mice and compare the survival of B cell subsets in wildtype vs. CD40 -/- or caspase-6 -/- mice. We predict that caspase-6 and CD40 are required for optimal B cell survival. We will also test if caspases selectively regulate homeostasis and expansion of B cell subsets. These studies will provide new insights into the mechanisms, which regulate the survival of normal B cells, malignant B cells and B cells contributing to autoimmune diseases